The secreted factor Sonic Hedgehog (Shh), which mediates signaling through three Gli transcription factors, is critical for organ development and proliferation of adult stem cells. Human mutations that reduce SHH signaling cause developmental defects, whereas inappropriate signaling causes cancer, likely through its action on stem cells. Promising therapies for such diseases involve the use of small molecule agonists and antagonists of the Shh pathway. To study the normal in vivo functions of each Gli transcription factor in Shh signaling, we generated an array of mouse mutants. A key finding of our work with direct relevance for translational research is Shh regulates the activity of each Gli in a distinct manner;Gli 1/2 as activators and GN3 as a represser. A second clinically relevant outcome is that the degree to which each Gli carries out Shh signaling is tissue specific. Therefore, in order to design therapies to treat diseases that result from altered Shh signaling, or augment stem cell activity, the contribution of Gli activator and represser function must be determined separately in each context. We recently devised an approach to mark and follow cells (including stem cells) responding to Shh signaling in vivo. An exciting finding of our work is that Shh regulates adult stem cells in diverse organs including forebrain neural stem cells and the stroma of epithelial organs. Furthermore, different Gli proteins act downstream of Shh in each population. We will uncover how Shh/Gli signaling regulates development of stem cell niches, and how Shh/Gli regulates stem cell behaviors in response to injury and cancer in the forebrain and prostate by: 1. Determining the roles of Shh signaling through each Gli in development of normal stem cell populations in the brain and prostate using fate mapping and conditional mutagenesis. 2. Determining the roles of Shh signaling through each Gli in regulating the response of forebrain neural stem cells and prostate stromal stem cells to injury, regeneration and cancer. Adult neural stem cells are stimulated to proliferate and migrate in response to brain tumors (gliomas) as well as to brain injury, and can have a positive therapeutic effect. On the other hand, neural stem cells have been implicated as the cell of origin of the most prevalent brain tumor, gliomas. As Shh/Gli signaling regulates neural stem cell proliferation, our studies will provide a rational basis for augmenting or reducing stem cell activity depending on the disease state. Likewise, in the prostate Shh is critical for expansion of the stroma during development, and has been implicated in the tumorigenesis of prostate cancer, the second most common cancer in men. Since mouse prostate stem cells can be manipulated in vivo through androgen driven regeneration following castration, this is an ideal system to determine how Shh/Gli signaling regulates stem cell populations in the prostate in normal and disease states.